The Characteristic Changes Of Circulating MicroRNA In The Clear Cell Renal Carcinoma Patients

Objective:To investigate the express of microRNA(miRNA) in the blood of clear cell renal cell carcinoma (ccRCC) patients and establish the distinctive circulating miRNA spectrum for the diagnosis and treatment of ccRCC.Methods:The total study population included 20 ccRCC patients and 20 control subjects. Among the ccRCC group there were 14 patients received radical nephrectomy,5 received nephron-spare hemi nephrectomy,1 received renal biopsy. All patients were diagnosed with pathology confirm. AJCC clinical stage included 8 in stageⅠ,3 in stageⅡ,5 in stageⅢand 4 in stageⅣ. The control group was consisted of healthy people. No diabetes in both groups. The blood samples were drawn with empty stomach. The blood samples were centrifugated. Plasma was aspirated and then frozen under the-80℃condition. Eight samples were chosen from ccRCC group and the control group. The plasma of those two groups was mixed with identical volume respectively. 2 ml plasma from each group was used to extract the total RNA by mirVana PARIS kit. After the saturation evaluation and quality control, the total RNA of each group was examed by TaqMan MicroRNA low density assay(TLDA) respectively. The result was analyzed through SDS 2.3 software and SPSS 15.0 software.Results:There were totally 83 miRNAs detected in both groups. There were 73 miRNAs detected in the ccRCC group and 55 detected in control group.28 miRNAs detected only in the ccRCC group and 10 detected only in the control group. According the judgment standard, we found 48 miRNAs up-regulated in ccRCC group and among them,24 were specific expressed. There were 12 miRNAs down-regulated in the ccRCC group contrasted to the control group of which 10 miRNAs were undetectable.Conclusion:[1] The circulating miRNA of ccRCC patients has its specific express spectrum relative to healthy people. [2] The expression changes of circulating miRNA in ccRCC patients mainly are up-regulation, those up-regulated miRNAs can distinct ccRCC patients from healthy people. We preliminarily establish the circulating miRNA express spectrum of ccRCC patients. Objective:To validate the result that miR-21, miR-142-5p and miR-34a were up-regulated in the plasma of ccRCC patients and analyze their relationship to the clinical stages of ccRCC.Methods:All of the samples in ccRCC group and control group were examed in this part. The total RNA of both ccRCC group and control group were extracted and tested respectively. Three of the up-regulated miRNAs, miR-21, miR-142-5p and miR-34a, were validated firstly using northern blot and then qRT-PCR analysis. All the samples of ccRCC group and control group were randomly paired. The miR-21, miR-142-5p and miR-34a of each sample in the ccRCC group were tested using qRT-PCR, the relationship of those miRNAs to the clinical stage of ccRCC were analyzed.Results:Northern blot result revealed that miR-21, miR-142-5p and miR-34a were up-regulated in the plasma of ccRCC patients which confirmed the results of miRNA assay results. qRT-PCR validation showed that the solubility curves of those three miRNA were good and no specific peak was found. The result also showed that miR-21, miR-142-5p and miR-34a were up-regulated in the plasma of ccRCC patients. They had very low expression in the control group. The qRT-PCR test of ccRCC patients showed that the up-regulation degree of miR-21 and miR-142-5p in the plasma had strong relation to the clinical stage. But miR-34a had no statistical significance between the up-regulation degree and the clinical stages of ccRCC.Conclusion:[1] The expression of circulating miR-21 and miR-142-5p in the ccRCC patients were up-regulated and the degree of up-regulation was strongly related to the clinical stages of ccRCC. [2]The expression of circulating miR-34a also was up-regulated in the ccRCC patients but its degree had no significant relation to the clinical stages. [3]Circulating miR-21, miR-142-5p and miR-34a could be used as tumor biomarkers for ccRCC.